The long term goal of this project is to determine the mechanisms by which bronchial artery perfusion prevents oxidant-mediated pulmonary vascular barrier dysfunction. The systemic bronchial circulation interacts with the pulmonary circulation by supplying vasa vasorum to the pulmonary arteries and veins and draining via bronchopulmonary anastomoses into the capillary and post-capillary pulmonary vascular bed. Based on our preliminary data in a pulmonary artery (PA) ischemia-reperfusion sheep lung preparation and in in vitro cocultures of sheep bronchial and PA endothelial cells, we hypothesize that perfusion of the vasa vasora of pulmonary vessels generates diffusible, protective mediators capable of countering the effects of reactive oxygen species on the pulmonary endothelium. These are likely nitric oxide (NO) and CO from heme oxygenase (HO). To test this hypothesis, we will first determine if bronchial artery flow protects via generation of NO by making direct measurements of NO and its metabolites. The effect of inhibiting bronchial circulation NOS followed by restoration of NO to the NOS-inhibited bronchial circulation with an NO infusion will be determined. In vitro studies will determine if bronchial microvascular endothelial cells protect pulmonary artery endothelial cells from oxidant-mediated injury and barrier dysfunction by an NO-dependent mechanism. Hydrogen peroxide-mediated endothelial NO production and Akt kinase-mediated endothelial NOS phosphorylation will be compared between cell types to explain the observed differences in response. We will examine the anatomy and physiology of the pulmonary vasa vasorum with fluorescent microspheres and determine the ability of PA vasa vasorum to generate NO and CO capable of diffusing into the lumen of an ischemic PA as a function of bronchial artery flow. Lastly, experiments in intact lungs and cultured cells will focus on potential down stream pathways mediating the protective effect of bronchial circulation NO on the pulmonary endothelium including CO and cGMP/protein kinase G. These results will provide new information about the physiology and function of pulmonary vasa vasorum and have significant implications with respect to clinical conditions where bronchial blood flow is reduced in the setting of an oxidant-mediated pulmonary insult such as lung transplantation.